title: Unique structural changes in calcium-bound calmodulin upon interaction with protein 4.1R FERM domain: novel insights into the calcium-dependent regulation of 4.1R FERM domain binding to membrane proteins by calmodulin.
abstract: Calmodulin (CaM) binds to the FERM domain of 80 kDa erythrocyte protein 4.1R (R30) independently of Ca(2+) but, paradoxically, regulates R30 binding to transmembrane proteins in a Ca(2+)-dependent manner. We have previously mapped a Ca(2+)-independent CaM-binding site, pep11 (A(264)KKLWKVCVEHHTFFR), in 4.1R FERM domain and demonstrated that CaM, when saturated by Ca(2+) (Ca(2+)/CaM), interacts simultaneously with pep11 and with Ser(185) in A(181)KKLSMYGVDLHKAKD (pep9), the binding affinity of Ca(2+)/CaM for pep9 increasing dramatically in the presence of pep11. Based on these findings, we hypothesized that pep11 induced key conformational changes in the Ca(2+)/CaM complex. By differential scanning calorimetry analysis, we established that the C-lobe of CaM was more stable when bound to pep11 either in the presence or absence of Ca(2+). Using nuclear magnetic resonance spectroscopy, we identified 8 residues in the N-lobe and 14 residues in the C-lobe of pep11 involved in interaction with CaM in both of presence and absence of Ca(2+). Lastly, Kratky plots, generated by small-angle X-ray scattering analysis, indicated that the pep11/Ca(2+)/CaM complex adopted a relaxed globular shape. We propose that these unique properties may account in part for the previously described Ca(2+)/CaM-dependent regulation of R30 binding to membrane proteins.
authors.affiliation: Department of Life Science, Graduate School of Engineering and Resource Science, Akita University, Tegata-Gakuên 1-1, Akita, 010-8502, Japan, nunomura@gipc.akita-u.ac.jp.
authors.forename: Wataru
authors.initials: W
authors.lastname: Nunomura
authors.forename: Noriyoshi
authors.initials: N
authors.lastname: Isozumi
authors.forename: Shigeyoshi
authors.initials: S
authors.lastname: Nakamura
authors.forename: Yuji
authors.initials: Y
authors.lastname: Jinbo
authors.forename: Shinya
authors.initials: S
authors.lastname: Ohki
authors.forename: Shun-Ichi
authors.initials: S
authors.lastname: Kidokoro
authors.forename: Hideki
authors.initials: H
authors.lastname: Wakui
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Takakuwa
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry* / Q000378:metabolism; D002413:Cations, Divalent; D002645:Chickens; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D056004:Molecular Dynamics Simulation; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010455:Peptides / Q000138:chemical synthesis / Q000737:chemistry*; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013816:Thermodynamics
chemical_list: D002147:Calmodulin; D002413:Cations, Divalent; D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D010455:Peptides; D011994:Recombinant Proteins; C028841:erythrocyte membrane band 4.1 protein; D002118:Calcium